Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Figure S1. Mating behavior of lep-2 mutants, graphical representation.
Each bar corresponds to an individual male. The amount of time it took for a male to contact a
hermaphrodite with his tail and to copulate is shown for each individual (spicule insertion was used to
identify copulation; sperm transfer was not assayed). If a male failed to sense or copulate within the
assay period of one hour, it was scored as “never”.
Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Figure S2. lep-2 mapping and genomic position
(A) Fluorescence ratios for probes of Chromosome IV. Raw data points are black dots, a
red line indicating smoothened average of data points and blue corresponds to the difference between the two values. Graph of probes containing the spike on the left arm of
Chromosome IV, showing the local linearity of the probe ratios. Red line indicates the
mean ratio for the left, middle and right segment.
(B) Genome browser-based view of the regions, including and flanking the 8 kb region
shown in panel A.
Figure S3. Expression of GFP::LEP-2
(A) Schematic of the transgene to express GFP::LEP-2, under the control of the lep-2 promoter.
(B,C) Bright field and fluorescence confocal micrographs of animals transformed with GFP::LEP-2.
(B) GFP::LEP-2 expression in the tail tip of a L3 and L4 stage male. (C) Representative images of
GFP::LEP-2 expression in various tissues and organs.
Development • Supplementary information
Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Development 143: doi:10.1242/dev.132738: Supplementary information
Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Figure S4. Expression of LIN-28::GFP in lep-2(bx73)
(A) DIC and fluorescence micrographs of tail tips in wild type and lep-2 mutant males
carrying the lin-28>lin-28::gf-p::lin-28_3’UTR transgene during larval development.
(B) Top: Quantification of Western blot data using the anti-LIN-28 antibody from
Weaver et al. (2014. eLife 3, p e04265). Values are normalized to levels in
wild-type L1 larvae. Actin was used as loading control. Bottom: Quantification of
Western blot data examining levels of LIN-28 in L3 in wild type and three lep-2
mutants using the anti-LIN-28 antibody from Seggerson et al. (2002. Dev. Biol. 243,
p. 215-225). Values are normalized to wild-type levels.
Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Movie 1. Mating behavior of wild-type C. elegans males
Development 143: doi:10.1242/dev.132738: Supplementary information
Development • Supplementary information
Movie 2. Mating behavior of lep-2 mutant C. elegans males
Development 143: doi:10.1242/dev.132738: Supplementary information
Table S1. Penetrance of delayed phenotypes in lep-2(lf) adults
Delayed phenotypes include molting adults and adults that are nonRol (rol-1 encodes an
adult-specific cuticle collagen).
n
52
46
17
20
15
24
34
n
289
258
125
% Molted
0
76
71
15
87
0
0
% nonRol
5
92
29
% Not molted
100
24
29
85
13
100
100
% Rol
95
8
71
Development • Supplementary information
Adult molts:
him-5(e1490)
lep-2(bx73); him-5
lep-2(bx147); him-5
lep-2(ok900); him-5
lep-2(sy68); him-5
him-5 post dauer
lep-2(bx73); him-5 post dauer
Adult Rol hermaphrodites:
rol-1(e91)
rol-1(e91); lep-2(ok900)
rol-1(e91); lep-2(ok900) post dauer
Development 143: doi:10.1242/dev.132738: Supplementary information
Table S2. Seam cell number and fusion of lateral alae are normal in lep-2 animals
Seam cell fusions were assayed using a GFP marker for adherens junctions, ajm-1::GFP;
seam cell nuclei were labeled with scm::GFP.
wIs78[ajm-1::gfp,
scm::GFP]
lep-2(ok900); wIs78
Avg. nr. of nuclei in the
seam1
50
Percent with fused
seam cells
16
100
40
16
n
100
Development • Supplementary information
Genotype
Development 143: doi:10.1242/dev.132738: Supplementary information
2
gfpnlp2_2
gaaaagttcttctcctttactcatttgctgaaaaatgtggaaaa
3
gfpnlp2_3
cacatttttcagcaaatgagtaaaggagaagaac
4
gfpnlp2_4
ctgtttcatgacgtggcattttgtatagttcatcca
5
gfpnlp2_5n
ggatgaactatacaaaatgccacgtcatgaaacagattgtcgat
6
lp2ex5r
ccaatcaataaatccttttcctgtccactttc
7
lp2ex5f
gagaacattttcgagaaaaatttgcgttttgg
8
gfpnlp2_6
caagcttcgccggatcaaatgacacac
9
lep2-7rn
ccgatttatcagcttcagctcaccgagagc
10
lep2-6f
cgacttacttcgctcaaatgccacaaaaattg
11
cln28dndraf
gaatagtaattcctctgatgaaatgaaccttattaaggaagatatgagag
12
ln283utrcdndrar
ctatcaatattctcagtgtctagatgattccatgcttgacttggtagaggactgtatcttgcaactg
13
ndndraln28r
gactctcatatcttccttaataaggttcatttcatcagaggaattactattcttttc
14
5plin28-1fnew
cagcattttcggtaaaactcttcaagcttg
15
cdndraln283utrf
16
3plin-28-6r
cagttgcaagatacagtcctctaccaagtcaagcatggaatcatctagacactgagaatattgata
g
gtattcaatcaatataaaaacaaaactctcg
17
plin28-7f
gtaaaactcttcaagcttgagggtg
18
3plin28-8r
cgatttatttcagcgttcgccc
19
lin-28_FW
tcgacggtagtatcggaggg
20
lin-28_RV
gaggtgttggtgacgggag
Development • Supplementary information
Table S3. Sequences of oligonucleotide primers used to make constructs.
Name
Sequence (5'–3')
1
gfpnlp2_1
ggccactcggtcctgttgaactgattt